Ionizing Radiation effects and Double-stranded breaks (DSBs) on the molecular karyotype of T. cruzi

• Grant number: 19/07916-7
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: June 01, 2019
• End date: July 31, 2020
• Field of knowledge: Biological Sciences - Parasitology - Protozoology of Parasites
• Principal Investigator: José Franco da Silveira Filho
• Grantee: Karina de Azevedo Vieira Honório
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
• Associated research grant: 16/15000-4 - Trypanosoma cruzi: intra and interspecific genomic variability and mechanisms of cell invasion/egress, AP.TEM

Abstract

Trypanosoma cruzi, the etiologic agent of Chagas disease, is a cell highly resistant to ionizing radiation. It can survive doses of gamma irradiation exposure that exceed considerably the absolutely lethal doses for mammalian cells. Lesions more complex and possibly more harmful to the cells are double-stranded breaks in DNA (DSBs), which can be repaired by homologous or nonhomologous recombination mechanisms. The DNA repair mechanisms may confer advantage to T. cruzi survival into mammalian host cells in which oxidative stress and the consequent formation of free radicals can cause lesions in the DNA. The exposure of this parasite to ionizing radiation produces intense chromosomal breakdown, visualized by the degradation of chromosomal bands shortly after irradiation. Then the fragmented genomic DNA is gradually reconstructed and the pattern of chromosomal bands is restored.We hypothesize that a small percentage of the population's cells are able to repair the DNA lesions and reconstruct the karyotype. To test this hypothesis, it is important to estimate the number of viable cells after irradiation necessary to ensure a resumption of growth. Regardless of the causes of DNA damage, cells activate defense mechanisms, such as cell cycle checkpoint arrest, apoptosis and DNA repair that can be detected and quantified.Epimastigotes from T. cruzi will be irradiated with doses of 100 and 500 Gy and maintained in axenic culture for 2-3 weeks, and the parameters mentioned above will be periodically determined (cell viability, cell cycle checkpoint arrest, apoptosis and DNA repair) and the molecular karyotype by PFGE. Thus we intend to obtain new information about the mechanisms involved in the reconstruction of parasite chromosomes.